Herpes simplex virus (HSV) keratitis is a leading cause of non-traumatic blindness in the US, with more than 200,000 cases per year. HSV can cause a variety of ocular diseases in humans ranging from self-limiting dendritic epithelial keratitis, conjunctivitis, and blepharitis to necrotizing stromal keratitis. In addition, HSV commonly causes cold sores, genital sores, and is a leading cause of viral encephalitis. The life cycles of HSV and other neurotropic herpesviruses are characterized by a lytic phase of infection at peripheral sites such as the cornea and skin during which all virus genes are expressed, and a latent phase of infection in neurons, during which gene expression is extremely limited. Latency represents a lifelong source of virus which can reactivate periodically causing severe ocular and other mucocutaneous damage. The unique regulatory switch between lytic and latent infection is poorly understood and the broad objectives of this proposal are therefore to elucidate this switch at the molecular level. This proposal aims to define viral genes which are critical for the establishment of, and reactivation from latency. To this end, mutations will be generated in cloned copies of HSV latency-related genes and the effects of these mutations tested in a series of in vitro assays. Selected mutations will then be introduced into the viral genome and recombinant viruses will be tested in a mouse ocular latency model. This approach will allow the identification of genes which impact the pathogenesis of HSV at the levels of establishment, maintenance, and reactivation of latency. A better understanding of the regulation of HSV gene expression in acutely and latently infected cells will allow further insight into the mechanisms by which HSV can persist for the lifetime of its host and indicate novel therapeutic approaches and targets for control of this blinding disease.